Isoproterenol-stimulated release of beta-endorphin and related peptides from the rat pituitary neurointermediate lobe in vitro: evidence for preferential release of certain molecular forms of beta-endorphin.
The intermediate lobe of the pituitary gland synthesizes the multifactorial precursor molecule pro-opiomelanocortin (POMC), from which, through a process of post-translational enzymatic processing, beta-endorphin-(1-31) (beta E) and a variety of N alpha-acetylated and C-terminally shortened forms of this peptide are generated. Using an in vitro superfusion system, the release of these endorphins from intact rat neurointermediate lobes (NILs) was investigated under basal and isoproterenol (ISO) stimulated conditions. Superfusion of NILs with the beta-adrenergic agonist ISO (30 min pulse) resulted in a rapid, sustained and concentration-dependent stimulation of the release of beta E-like immunoreactivity (beta E-IR) over basal as determined with an antiserum directed against the C-terminus of the beta E- (1-31) sequence (10(-6) M: + 145%; 10(-7) M: + 73%; 10(-8) m: + 41%). The release of N(alpha)-acetylated-endorphin-like immunoreactivity (AcE-IR) was stimulated to a similar extent. These effects of ISO were antagonized by the competitive alpha-adrenoceptor antagonist propranolol in a concentration-dependent manner, indicating the involvement of alpha-adrenoceptors. The beta-related peptides released from the NILs under basal and ISO-stimulated conditions were further characterized, based on their retention times in a reversed-phase HPLC system and their reactivity with specific antisera recognizing respectively the midportion of beta E, the N-terminus of acetylated endorphins, the C-terminus of tau-endorphin (beta E-(1-17); tau E), or the C-terminus of alpha-endorphin (beta E-(1-16); alpha E). In HPLC fractionated superfusates 10 peaks were resolved that reacted with the midportion beta E antiserum. In superfusates collected under basal conditions, three major peaks possessed chromatographical and immunological characteristics of Ac beta E-(1-26), Ac beta E- (1-27) Ac beta E-(1-31). In addition, a prominent peak was found eluting around the retention time of beta E-(1-31), that contained both acetylated and non-acetylated material. Six smaller peaks were observed, with the characteristics of beta E-(1-26) and beta E-(1-27) (these peptides were not resolved with the HPLC system used), Ac tau E, tau E, Aa alpha E, and des-tyrosine-alpha E (DT alpha E), respectively. In superfusates collected during superfusion of NILs with ISO (10(-6) M) all peaks were increased. However, those eluting as beta E-(1-31), beta E-(1-26)/beta E-(1-27), Ac beta E-(1-26) and Ac tau E appeared to be preferentially stimulated.(ABSTRACT TRUNCATED AT 400 WORDS)
Sweep
C G
CG
Rudolf Magnus Institute, Department of Pharmacology, Medical Faculty, University of Utrecht, The Netherlands.
Boersma
C J
CJ
Wiegant
V M
VM
D000107:Acetylation; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D007545:Isoproterenol / Q000494:pharmacology*; D007700:Kinetics; D008297:Male; D010904:Pituitary Gland, Posterior / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D001615:beta-Endorphin / Q000302:isolation & purification / Q000378:metabolism*
D001615:beta-Endorphin; D007545:Isoproterenol